Child Melioidosis Deaths Caused by Burkholderia pseudomallei-Contaminated Borehole Water, Vietnam, 2019.

Within 8 months, 3 children from 1 family in northern Vietnam died from melioidosis. Burkholderia pseudomallei of the same sequence type, 541, was isolated from clinical samples, borehole water, and garden and rice field soil. Boreholes should be properly constructed and maintained to avoid B. pseudomallei contamination.

Clinical Utility of Combined Whole-cell Antigen and Recombinant Hemolysis Co-regulated Protein 1-Enzyme-linked Immunosorbent Assays Reveals Underdiagnosed Cases of Melioidosis in Vietnam.

Melioidosis is a fatal infectious disease in the tropics and subtropics. Currently, bacterial culture is the gold standard for diagnosis of the disease, but its sensitivity is relatively low. In this study, we evaluated four ELISAs using sera collected from culture-confirmed cases of melioidosis (n = 63), cases with other bacterial infections (n = 62), and healthy donors (n = 60). Antigens used for ELISAs were the whole-cell (WC) antigens and recombinant proteins of hemolysis co-regulated protein 1 (Hcp1), GroEL1, and alkyl hydroperoxide reductase subunit C (AhpC). Using the cutoff values for optical density at 490 nm defined at a specificity of > 95%, the sensitivity of the WC, Hcp1, GroEL1, and AhpC ELISAs was 93.7%, 87.3%, 61.9%, and 57.1%, respectively. The combined WC/Hcp1 ELISA showed the greatest sensitivity and specificity of 98.4% and 95.1%, respectively. Of 511 and 500 sera collected from clinically suspected febrile patients admitted to the General Hospital of Ha Tinh Province and the Hue Central Hospital, respectively, combined WC/Hcp1 ELISAs showed 52 (10.2%) and 41 (8.2%) patients positive for melioidosis, respectively. The assay detected 14 of 14 (100%) and 21 of 23 (91.3%) culture-confirmed cases of melioidosis at Ha Tinh and Hue, respectively. A follow-up study of 38 patients positive for melioidosis by combined WC/Hcp1 ELISAs but negative for Burkholderia pseudomallei by culture method or not assigned to examine for bacterial culture resulted in 2 (5.3%) culture-reconfirmed patients with melioidosis, 9 (23.7%) deaths, 17 (44.7%) unhealthy patients, and 10 (26.3%) healthy persons. Combined WC/Hcp1 ELISA was a reliable serological method to detect underdiagnosed cases of melioidosis. Further investigations are needed to estimate the true sensitivity and specificity of the assay and the true number of cases of melioidosis.

Erythritol as a single carbon source improves cultural isolation of Burkholderia pseudomallei from rice paddy soils.

BACKGROUND: Isolation of the soil bacterium Burkholderia pseudomallei from tropical environments is important to generate a global risk map for man and animals to acquire the infectious disease melioidosis. There is increasing evidence, that the currently recommended soil culture protocol using threonine-basal salt solution with colistin (TBSS-C50) for enrichment of B. pseudomallei and Ashdown agar for subsequent subculture lacks sensitivity. We therefore investigated, if the otherwise rarely encountered erythritol catabolism of B. pseudomallei might be exploited to improve isolation of this bacterium from soil. METHODOLOGY/PRINCIPAL FINDINGS: Based on TBSS-C50, we designed a new colistin-containing medium with erythritol as the single carbon source (EM). This medium was validated in various culture protocols by analyzing 80 soil samples from 16 different rice fields in Vietnam. B. pseudomallei enrichment was determined in all culture supernatants by a specific quantitative PCR (qPCR) targeting the type three secretion system 1. 51 out of 80 (63.8%) soil samples gave a positive qPCR signal in at least one of the culture conditions. We observed a significantly higher enrichment shown by lower median cycle threshold values for B. pseudomallei in a two-step culture with TBSS-C50 for 48 h followed by EM for 96h compared to single cultures in TBSS-C50 for either 48h or 144h (p<0.0001, respectively). Accordingly, B. pseudomallei could be isolated on Ashdown agar in 58.8% (30/51) of samples after subcultures from our novel two-step enrichment culture compared to only 9.8% (5/51) after standard enrichment with TBSS-C50 for 48h (p<0.0001) or 25.5% (13/51; p<0.01) after TBSS-C50 for 144h. CONCLUSIONS/SIGNIFICANCE: In the present study, we show that specific exploitation of B. pseudomallei metabolic capabilities in enrichment protocols leads to a significantly improved isolation rate of this pathogen from soil compared to established standard procedures. Our new culture method might help to facilitate the creation of environmental risk maps for melioidosis in the future.